Galactokinase deficiency is an autosomal recessive disorder that causes cataracts in humans. Homozygotes develop cataracts within the first year of life; heterozygotes develop cataracts at a later age. The long term objectives of this proposal are to characterize the mutations in the human population which are associated with cataracts and galactokinase deficiency, produce an animal model of galactokinase deficiency and finally, identify the best mode of gene therapy for hereditary cataracts using galactokinase as a model. As a starting point for these long term objectives, we propose to first characterize the genomic structure of human galactokinase. Knowledge of the genomic structure will allow characterization of mutations on patient DNA. Next we will isolate the mouse galactokinase gene in order to insert this gene (after designing it to produce a null mutation) into mouse blastocysts to produce a transgenic mouse with galactokinase deficiency. The galactokinase deficient mouse will serve the scientific community as a galactosemic animal model to study a)galactose metabolism, b)cataract, retinopathy and peripheral neuropathy, c)efficacy of aldose reductase inhibitors, and d)as an animal model to study gene therapy of galactosemia. Finally, we will subclone the human galactokinase gene into an adenoviral vector to determine the best mode of gene therapy for this disorder. We propose to insert the normal gene into tissue culture cells for our initial experiments. Next, in vivo experiments will be performed to determine the best mode of delivery and dosage for gene delivery to the lens. First, an adenovirus containing lacZ will be delivered to a normal mouse lens to determine the appropriate titer and most efficient route of delivery. Following these initial experiments, an adenovirus containing the galactokinase gene will be delivered to the lens of the galactokinase deficient mouse in order to correct the enzyme defect. The information obtained from the proposed experiments will not only improve the management of galactokinase deficiency but provide insight into the gene therapy for hereditary cataracts.